This project is designed to determine the pathogenesis of hemolysis-induced pigment gallstone disease using the laboratory mouse (genotype nb/nb) with hereditary hemolytic anemia as an animal model of the human disease. Although hemolysis and cirrhosis are closely associated with pigment gallstones, patients without those conditions have pigment stone disease and account for about 1/3 of the 500,000 cholecystectomies performed yearly. Since the etiology of pigment gallstones is unknown, the long-term goal of this project is to use the mouse model to better understand the determinants of hemolysis-related gallstone disease and to develop methods of prevention and dissolution. specifically, we will (1) determine the solubility of unconjugated bilirubin in model bile solutions in the presence or absence of calcium; (2) construct a miniature calcium ion selective membrane electrode to directly measure calcium activity in model solutions; (3) determine the calcium activity and unconjugated bilirubin concentrations in bile of nb/nb mice with or without stones and the solubility product of calcium bilirubinate in mouse bile in vitro; (4) determine the role of the anemia independent of otyher genetic factors by transplantation of nb/nb bone marrow into another genotype W/W v and subsequent analysis of bile composition; (5) determine the effect of increased loads of bilirubin and bile salts on calcium activity and unconjugated bilirubin concentration in bile of mice with or without stones; (6) determine the role of hydrolysis of conjugated to unconjugated bilirubin in bile; and (7) assess gallbladder contractility in vitro by isolated tissue bath and the dose response to the octapeptide of cholecystokinin and the effect of unconjugated bilirubin, calcium activity and estrogens on that response. These studies intertwine the disciplines of physical chemistry, genetics and smooth muscle physiology.